Transcription mechanisms that contribute to HIV-1 latency

描述(申请人提供)：HIV前病毒在转录水平上受到调控，涉及转录起始、染色质组织和延伸的变化。特别是，转录延长已被证明是HIV表达的限制步骤。我们假设，RNA聚合酶II的处理能力和提前转录终止与染色质重塑的协调控制创建了一个关键检查点，限制了前病毒转录，并直接影响不同细胞和分子环境中的HIV潜伏期。我们已经证明，RNA聚合酶II的处理能力是HIV转录的检查点，而NELF是一种暂停RNA聚合酶II的因子，它通过招募终止因子Pcf11和HIV-1长端重复序列的转录辅阻遏物来重新抑制HIV前病毒。利用多种生化方法，包括染色质免疫沉淀、RNAPII足迹、体外转录系统和质谱学，我们建议确定负调控HIV转录延伸的生化机制和表征细胞因素。这些研究将为艾滋病毒潜伏期的建立、维持和逆转提供新的见解。了解HIV转录延长的调控将为控制和清除不同细胞库中的HIV提供新的细胞靶点。

DESCRIPTION (provided by applicant): HIV provirus is regulated at the level of transcription and involves changes in transcription initiation, chromatin organization and elongation. In particular, transcription elongation has been demonstrated to be a limiting step for HIV expression. We hypothesize that the coordinated control of RNA polymerase II processivity and premature transcription termination coupled with chromatin remodeling creates a key checkpoint that limits provirus transcription and directly impact HIV latency in different cellular and molecular contexts. We have shown that RNA polymerase II processivity is a check point for HIV transcription and that NELF, a factor that pauses RNA polymerase II, re-enforces repression of HIV proviruses by recruiting a termination factor Pcf11 and transcriptional corepressors to the HIV-1 long terminal repeat. Using a variety of biochemical approaches, including chromatin immunoprecipitation, RNAPII footprinting, in vitro transcription systems and mass spectrometry, we propose to determine the biochemical mechanisms and characterize the cellular factors that negatively regulate HIV transcription elongation. These studies will provide new insights into the establishment, maintenance and reversal of HIV latency. Understanding the regulation of HIV transcription elongation will provide novel cellular targets for controlling and purging HIV in different cellular reservoirs.